Development of a vaccine for respiratory syncytial virus (RSV) has received a high priority. The major impediment to initiating clinical trials with new candidate vaccines is an incomplete understanding of the vaccine-enhanced illness caused by formalin-inactivated RSV vaccines. Our group has described a BALB/c mouse model of RSV in which details of the immunologic mechanisms of RSV vaccine-immunopotentiation can be studied. We have shown that priming mice with inactivated RSV antigen induces a Th2-like cytokine mRNA expression pattern after RSV challenge (dominant IL-4 expression), and that live RSV priming by the nasal or parenteral route induces a Th1-like cytokine mRNA expression pattern after RSV challenge (dominant IFN-gamma expression). This finding has led us to propose the hypothesis that selective activation of the Th2 cell subset is responsible for RSV vaccine immunopotentiation of disease. The theme of this proposal is to address the impact of the cytokine milieu on the pattern of immune response to RSV immunization and challenge in both mice and humans. The specific aims are to: l) define the influence of the local and systemic cytokine environment on the response to RSV immunization and challenge in mice; 2) determine whether antigen specificity or mechanism of antigen presentation is the major factor in the selective activation of T cell subsets and pattern of cytokine expression in RSV-infected mice; and 3) test the validity of the mouse model by evaluating cytokine expression patterns in RSV-infected children. Aims l and 2 will be approached by using transgenic mice with "knock-outs" in specific cytokines, and recombinant vaccinia vectors that cc-express individual RSV proteins and specific murine cytokines. The human studies will take advantage of a clinical trial infrastructure for RSV therapeutics, and ongoing vaccine trials of live attenuated RSV to obtain serum, PBMC, nasal, and tracheal secretions for cytokine analysis. The assay detection methods for the human studies will include EIA for direct cytokine measurement, and quantitative RNA PCR for cytokine message. Validating the mouse as a model for evaluating patterns of immune response to RSV will accelerate preclinical development of candidate RSV vaccines. Defining how the cytokine milieu influences the immune response to RSV vaccination and challenge will impact strategies for RSV vaccine design and delivery.